The cII protein of bacteriophage lambda is a key regulatory protein in the life cycle of the virus. If sufficiently high levels of cII protein are present, the virus enters the lysogenic pathway and the phage DNA is inserted into the host DNA as a dormant prophage; if not enough cII protein is present, the virus enters the lytic pathway and progeny virus particles are released with lysis of the host. The level of cII protein itself is dependent upon the host and viral genotype, and also upon various environmental factors. Under normal conditions all of these factors are exquisitely balanced so that some infected cells enter the lytic pathway and others the lysogenic pathway. The long term objective of this research is to understand the interplay of all of these factors, not only because this is an important problem in its own right, but also because the understanding of this system promises to yield important insights into the regulation of basic cellular processes. A specific aim for this grant period is to understand the role of RNA processing in the modulation of cII gene activity, including the alternate pathway for processing of cII mRNA that is utilized in the absence of the OOP antisense RNA, and a second alternate processing pathway, dependent upon the OOP antisense RNA, that is utilized in the absence of the host enzyme RNase III. In order to identify additional host genes that influence cII gene activity, a library of 8 kb E. coli DNA fragments under control of the strong lambda pL promoter has been screened for clones that inhibit lambda cII gene activity. A second specific aim is to investigate the nature and mode of action of genes which, when overexpressed, inhibit lambda cII gene activity. A third specific aim concerns the comparative biology of cII action in the lambdoid phages lambda, 21, P22 and phi80 on the cII-dependent pRE and pI promoters in these phages. (Phage phi80 has no pRE promoter, and cII- dependent repressor transcription occurs from the phi80 pRM promoter. In lambda, pI governs synthesis of lambda int mRNA; in P22 pI is headed in the wrong direction and may result in synthesis of an antisense RNA that results in processing about 100 nucleotides away from the region of complementarity; and in phi80 the int gene is pointed in the reverse direction with respect to the other phages, with nothing known about a possible pI promoter.) The nature of these systems and the mechanisms by which they lead to regulation of int mRNA levels will be investigated.